Characterization Of Arginase From The Liver Of Mango Tilapia (Sarotherodongalilaeus)

Project and Seminar Material for Biochemistry

Characterization Of Arginase From The Liver Of Mango Tilapia (Sarotherodongalilaeus)


Arginase is a detoxifying enzyme that catalyses the hydrolysis of arginine into ornithine and urea, the last step of urea cycle- a process through which the body disposes off harmful ammonia. This research was carried out to determine the characteristics of liver arginase of mango tilapia (Sarotherodongalilaeus) in Opa river, Osun state.

The enzyme ‘arginase’ was isolated from the liver of mango tilapia through the process of homogenization and centrifugation which was done at 4000rev/min.The protein concentration was determined using Bradford method and the arginase assay was determined by Kaysen and Strecker method.

The kinetic study shows that mango tilapia of liver arginase has a Km value of 0.2M and a Vmax value of 166.7µmol/ml/min. The effect of temperature on arginase activity was tested and the optimum temperature for mango tilapia liver arginase is 50°C at activity of 63.44µmol/ml/min. The effect of pH was also investigated and optimum pH is 8.0 at activity of 165.1µmol/ml/min. Inhibition study was also carried out and it was observed that calcium (51.4±1.13) and zinc (51.5±4.27) strongly inhibit arginase while mercury, magnesium and sodium have little or no inhibitory effect on arginase. Also from the result, it can be deduce that citrate (20.8±4.67) and glutathione (28.6±2.53) and ethylenediamineacetic acid (EDTA) slightly inhibit arginase while urea has les inhibitory effect on arginase. The result for the effect of amino acids on enzyme activity shows that liver arginase of mango tilapia to be in this order: arginine > valine> aspartate > cysteine > lysine with residual activity of 107.2%, 86.6%, 73.4%, 62.1% and 58.7% respectively.

Mango tilapia liver arginase belongs to the ureotelic class of arginases according to Mora J et al classification (1965). It has a Km value of 0.2M and aVmax value of 166.7µmol/ml/min.

Table Of Contents

  • Title page i
  • Certification page ii
  • Dedication iii
  • Acknowledgement iv
  • Abstract v
  • Table of content

Chapter One:

  • Introduction And Literature Review
  • 1.o Introduction
  • 1.2 Classification
  • 1.3 Distribution and occurrence
  • 1.4 Literature review
  • 1.4.1 Arginase
  • 1.4.2 Arginase isoenzymes
  • 1.5 Structural properties
  • 1.5.1 Molecular properties
  • 1.5.2 Amino acid composition
  • 1.6 Physicochemical properties
  • 1.6.1 Kinetic properties
  • 1.6.2 Effect of p H
  • 1.6.3 Effect of temperature
  • 1.6.4 Effect of metal ions
  • 1.7 Mechanism of action
  • 1.8 Arginase deficiency
  • 1.9 Biological and physiological importance of arginase
  • 1.10 Study sample
  • 1.10.1 Taxonomy
  • 1.10.2 Morphology
  • 1.10.3 Distribution
  • 1.10.4 Ecology
  • 1.10.5 Biology
  • 1.10.6 Feeding habits
  • 1.10.7 Reproduction and parental care
  • 1.10.8 Economic importance
  • 1.11 Liver
  • 1.12 Objectives

Chapter Two:

  • Materials And Methods
  • 2.1 Materials
  • 2.1.1 Chemicals and reagents
  • 2.1.2 Equipments
  • 2.2 Methods
  • 2.2.1 Enzyme isolation
  • 2.2.2 Preparation of buffer and reagents
  • 0.33M Arginine
  • Bradford reagent
  • Erlich reagent
  • Homogenization buffer (0.1M Phosphate buffer)
  • 0.1M Citrate buffer, p H 3.0
  • 0.1M Trizma HCl buffer, p H 6.0
  • 0.1M Trizma HCl buffer, p H 7.0
  • 0.1M Trizma HCl buffer, p H 9.0
  • Arginase assay
  • Protein assay
  • Determination of kinetic parameters
  • Effect of p H on enzyme activity
  • Effect of temperature on enzyme activity
  • Effect of amino acids on enzyme activity
  • Effect of chelating compounds on arginase activity
  • Effect of metal ions on arginase activity

Chapter Three:

  • Results
  • 3.1 Effect of temperature
  • 3.2 Effect of pH
  • 3.3 Kinetic study
  • 3.4 Inhibition study
  • 3.4.1 Effect of chelating compounds
  • 3.4.2 Effect of divalent metals
  • 3.4.3 Effect of amino acids

Chapter Four:

  • Discussion, Conclusion And Recommendation
  • 4.1 Discussion
  • 4.2 Conclusion
  • 4.3 Recommendation
  • References

List Of Tables

  • Table 3.1 Effect of temperature on arginase activity
  • Table 3.2 Effect of pH on arginase activity
  • Table 3.3 Kinetic study
  • Table 3.4 Effect of chelating compounds
  • Table 3.5 Effect of divalent metals
  • Table 3.6 Effect of amino acids

List Of Figures

  • Figure 1.1 External structure of mango tilapia
  • Figure 3.1 Graph of temperature against activity
  • Figure 3.2 Graph of pH against activity
  • Figure 3.3 Determination of Km and Vmax

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